Dataset Information

Mitochondrial uncoupler carbonyl cyanide m-chlorophenylhydrazone induces vasorelaxation without involving K_ATP channel activation in smooth muscle cells of arteries.

ABSTRACT:

Background and purpose

The effects and mechanisms of chemical mitochondrial uncouplers on vascular function have never been identified. Here, we characterized the effects of the typical mitochondrial uncoupler carbonyl cyanide m-chlorophenylhydrazone (CCCP) on vascular function in rat mesenteric arteries and aorta and elucidated the potential mechanisms.

Experimental approach

Isometric tension of mesenteric artery and thoracic aorta was recorded by using a multiwire myograph system. Protein levels were measured by western blot analyses. Cytosolic [Ca²⁺ ]_i , mitochondrial ROS (mitoROS) and mitochondrial membrane potential of smooth muscle cells (A10) were measured by laser scanning confocal microscopy.

Key results

Acute treatment with CCCP relaxed phenylephrine (PE)- and high K⁺ (KPSS)-induced constriction of rat mesenteric arteries with intact and denuded endothelium. Pretreatment with CCCP prevented PE- and KPSS-induced constriction of rat mesenteric arteries with intact and denuded endothelium. Similarly, CCCP prevented PE- and KPSS-induced constriction of rat thoracic aorta. CCCP increased the cellular ADP/ATP ratio in vascular smooth muscle cells (A10) and activated AMPK in A10 cells and rat thoracic aorta tissues. CCCP-induced aorta relaxation was attenuated in AMPK ?1 knockout (-/-) mice. SERCA inhibitors thapsigargin and cyclopiazonic acid (CPA) but not the K_ATP channel blocker glibenclamide partially inhibited CCCP-induced vasorelaxation in endothelium-denuded rat mesenteric arteries. CCCP increased cytosolic [Ca²⁺ ]_i , mitoROS production and depolarized mitochondrial membrane potential in A10 cells. FCCP, the analogue of CCCP, had similar vasoactivity as CCCP in rat mesenteric arteries.

Conclusions and implications

CCCP induces vasorelaxation by a mechanism that does not involve K_ATP channel activation in smooth muscle cells of arteries.

SUBMITTER: Zhang YQ

PROVIDER: S-EPMC5056225 | biostudies-literature | 2016 Nov

REPOSITORIES: biostudies-literature

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Publications

Mitochondrial uncoupler carbonyl cyanide m-chlorophenylhydrazone induces vasorelaxation without involving K<sub>ATP</sub> channel activation in smooth muscle cells of arteries.

Zhang Yan-Qiu YQ Shen Xin X Xiao Xiao-Lin XL Liu Ming-Yu MY Li Shan-Liang SL Yan Jie J Jin Jing J Gao Jin-Lai JL Zhen Chang-Lin CL Hu Nan N Zhang Xin-Zi XZ Tai Yu Y Zhang Liang-Shuan LS Bai Yun-Long YL Dong De-Li DL

British journal of pharmacology 20160909 21

<h4>Background and purpose</h4>The effects and mechanisms of chemical mitochondrial uncouplers on vascular function have never been identified. Here, we characterized the effects of the typical mitochondrial uncoupler carbonyl cyanide m-chlorophenylhydrazone (CCCP) on vascular function in rat mesenteric arteries and aorta and elucidated the potential mechanisms.<h4>Experimental approach</h4>Isometric tension of mesenteric artery and thoracic aorta was recorded by using a multiwire myograph syste ...[more]

PMID: 27534899

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Project description:Experimental evolution of Escherichia coli K-12 with benzoate, a partial uncoupler of the proton motive force (PMF), selects for mutations that decrease antibiotic resistance. We conducted experimental evolution in the presence of carbonyl cyanide m-chlorophenylhydrazone (CCCP), a strong uncoupler. Cultures were serially diluted daily 1:100 in LBK medium containing 20 to 150?µM CCCP buffered at pH 6.5 or at pH 8.0. After 1,000 generations, the populations tolerated up to 150?µM CCCP. Sequenced isolates had mutations in mprA (emrR), which downregulates the EmrAB-TolC pump that exports CCCP. A mprA::kanR deletion conferred growth at 60 ?M CCCP, though not at the higher levels resisted by evolved strains (150?µM). Some mprA mutant strains also had point mutations affecting emrA, but deletion of emrA abolished the CCCP resistance. Thus, CCCP-evolved isolates contained additional adaptations. One isolate lacked emrA or mprA mutations but had mutations in cecR (ybiH), whose product upregulates drug pumps YbhG and YbhFSR, and in gadE, which upregulates the multidrug pump MdtEF. A cecR::kanR deletion conferred partial resistance to CCCP. Other multidrug efflux genes that had mutations included ybhR and acrAB The acrB isolate was sensitive to the AcrAB substrates chloramphenicol and tetracycline. Other mutant genes in CCCP-evolved strains include rng (RNase G) and cyaA (adenylate cyclase). Overall, experimental evolution revealed a CCCP-dependent fitness advantage for mutations increasing CCCP efflux via EmrA and for mutations that may deactivate proton-driven pumps for drugs not present (cecR, gadE, acrAB, and ybhR). These results are consistent with our previous report of drug sensitivity associated with evolved benzoate tolerance.IMPORTANCE The genetic responses of bacteria to depletion of proton motive force (PMF), and their effects on drug resistance, are poorly understood. PMF drives export of many antibiotics, but the energy cost may decrease fitness when antibiotics are absent. Our evolution experiment reveals genetic mechanisms of adaptation to the PMF uncoupler CCCP, including selection for increased CCCP efflux but also against the expression of PMF-driven pumps for drugs not present. The results have implications for our understanding of the gut microbiome, which experiences high levels of organic acids that decrease PMF.

Project description:Obesity plays an important role in the pathogenesis of hypertension. Renal dopamine D1-like receptor-mediated diuresis and natriuresis are impaired in the obese Zucker rat, an obesity-related hypertensive rat model. The role of arterial D1 receptors in the hypertension of obese Zucker rats is not clear.Plasma glucose and insulin concentrations and blood pressure were measured. The vasodilatory response of isolated mesenteric arteries was evaluated using a small vessel myograph. The expression and phosphorylation of D1 receptors were quantified by co-immunoprecipitation and immunoblotting To determine the effect of hyperinsulinemia and hyperglycemia on the function of the arterial D1 receptor, we studied obese Zucker rats (six to eight-weeks old) fed (6 weeks) vehicle or rosiglitazone, an insulin sensitizer (10 mg/kg per day) and lean Zucker rats (eight to ten-weeks old), fed high-fat diet to induce hyperinsulinemia or injected intraperitoneally with streptomycin (STZ) to induce hyperglycemia.In obese Zucker rats, the vasorelaxant effect of D1-like receptors was impaired that could be ascribed to decreased arterial D1 receptor expression and increased D1 receptor phosphorylation. In these obese rats, rosiglitazone normalized the arterial D1 receptor expression and phosphorylation and improved the D1-like receptor-mediated vasorelaxation. We also found that D1 receptor-dependent vasorelaxation was decreased in lean Zucker rats with hyperinsulinemia or hyperglycemia but the D1 receptor dysfunction was greater in the former than in the latter group. The ability of insulin and glucose to decrease D1 receptor expression and increase its phosphorylation were confirmed in studies of rat aortic smooth muscle cells.Both hyperinsulinemia and hyperglycemia caused D1 receptor dysfunction by decreasing arterial D1 receptor expression and increasing D1 receptor phosphorylation. Impaired D1 receptor-mediated vasorelaxation is involved in the pathogenesis of obesity-related hypertension.

Dataset Information

Mitochondrial uncoupler carbonyl cyanide m-chlorophenylhydrazone induces vasorelaxation without involving K_ATP channel activation in smooth muscle cells of arteries.

Background and purpose

Experimental approach

Key results

Conclusions and implications

Publications

Mitochondrial uncoupler carbonyl cyanide m-chlorophenylhydrazone induces vasorelaxation without involving K<sub>ATP</sub> channel activation in smooth muscle cells of arteries.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets

Dataset Information

Mitochondrial uncoupler carbonyl cyanide m-chlorophenylhydrazone induces vasorelaxation without involving KATP channel activation in smooth muscle cells of arteries.

Background and purpose

Experimental approach

Key results

Conclusions and implications

Publications

Mitochondrial uncoupler carbonyl cyanide m-chlorophenylhydrazone induces vasorelaxation without involving K<sub>ATP</sub> channel activation in smooth muscle cells of arteries.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets

Mitochondrial uncoupler carbonyl cyanide m-chlorophenylhydrazone induces vasorelaxation without involving K_ATP channel activation in smooth muscle cells of arteries.